Many fields of science and medicine require the close observation of small samples, for example, thin tissue slices extracted from living organisms or living/fixed cell cultures. Currently, the leading modalities for these kinds of observations are optical and fluorescence microscopy. These techniques are very mature and provide rich information regarding the investigated sample. Nevertheless, such optical-based modalities lack the ability to observe moderately thick three-dimensional (3D) non-transparent samples. They can not measure vital parameters such as molecular self diffusion and 3D flow vectors, and, for example in terms of medical tissues, many times result in inconclusive clinical diagnosis of histological samples. Furthermore, using optical methods, the following difficulties are encountered: it is difficult to image accurately the O2 partial pressure in specimens; it is difficult to recognize with high specificity various superoxides in the imaged sample; one can not measure a variety of image contrasts such as the spatially-resolved magnetic resonance relaxation times (T1 and T2) and the lineshape of the spins in the sample; and optical methods lack the possibility to correlate in-vitro with in-vivo measurements.
Nuclear Magnetic Resonance (NMR) microscopy, in which the nuclear spins can be considered as a unique kind of magnetic stain or dye, is currently a relatively widespread complimentary imaging tool for small samples that is found useful in many diverse clinical situations to achieve high identification specificity and measurement accuracy in cases of inflammation, fluid diffusion, blood flow and perfusion, lipid content, tissue types such as cancer, and tissue necrosis. NMR microscopy has also been found useful in materials science and botany to investigate and measure flow and porosity. Several NMR devices and applications of NMR microscopy are described in U.S. Pat. Nos. 5,258,710, 5,394,088, and 5,416,414. Thus, NMR microscopes are routinely employed in many fields of science and medicine, and several companies are producing such instruments commercially. Some people have even combined NMR and optical microscopes. The main drawback of the existing NMR microscopes is their high price (in the range of $500,000 to $1,500,000, mainly due to the superconducting magnet technology required for their operation), and the limited image resolution they offer (>10 microns), which can not rival the <0.5 micron resolution of optical imaging modalities. Due to these limitations (and regardless of the many potential advantages), the full potential of magnetic resonance microscopy cannot be readily exploited and such instruments are presently much less abundant than the optical-based microscopes in scientific and medical laboratories around the world.
Another, less common technique of magnetic resonance imaging, employs electron spin resonance (ESR) in paramagnetic molecules, rather than the spins of hydrogen nuclei. Whereas the field of NMR microscopy is well developed, ESR microscopy is not. Nevertheless, ESR has inherently many potential virtues over NMR, which could make this a technique of choice for Magnetic Resonance (MR) microscopic applications. For example, the signal per spin is much higher than in NMR, diffusion does not limit the resolution in the short time scales (T2's˜μs) of the ESR measurements, ESR micro-resonators detect with a quality factor (Q) of ˜1000 compared to the Q˜10 of the NMR micro-coils, and the ESR lineshape is more sensitive to dynamic effects, leading to richer information. An additional factor is the low cost of electromagnets used in ESR as compared to the expensive superconducting magnets of NMR microscopes. Since most samples do not contain stable paramagnetic molecules, paramagnetic species (often in the form of stable organic radicals) must be added in a manner similar to that of adding contrast agents in NMR or dyes in optics. This is a standard procedure, especially for microscopy, which also offers the benefit of eliminating any concerns associated with a large undesirable background signal, (such as protons in NMR). An ESR microscope can provide similar spatially resolved sample parameters to those obtained by NMR measurements, (i.e. spin concentration, relaxation times T1, T2, and diffusion coefficient), which compliments the information obtained by conventional optical microscopy.
Up to now, most ESR imaging (ESRI) efforts in biological samples have been directed towards observation of large subjects and to determining the radical and oxygen concentration (by its effect on the radical line width). U.S. Pat. Nos. 5,502,386, 5,578,922, 5,678,548 and 5,865,746 describe some of these efforts. Such experiments, conducted in-vivo, employ low fields of ˜10 mT at low RF frequencies (which results in relatively low spin sensitivity), so that the RF energy will penetrate deeply into the relatively large biological object. Consequently, a typical voxel resolution in low frequency ESR experiments is ca. [2 mm]3. Most low-field ESR imaging techniques are based on Continuous Wave (CW) detection where the image is obtained by applying static gradients in various directions with respect to the object, which is sometimes referred to as the back projection technique. However, utilization of a single pulse Free Induction Decay (FID) sequence in conjunction with pulsed and static gradients has been also explored.
Previous publications, including a paper from one the inventors, have described in the past an ESR resonator based on high permittivity KTaO3 and demonstrated its application in the field of ESR spectroscopy. Such high permittivity small resonator structures, however, were not employed in the past as a basis for a miniature ESR imaging probe (which includes a resonator and the imaging gradient coils).
Other emerging techniques of high resolution ESR imaging include the use of magnetic tips (Magnetic Resonance Force Microscopy), Hall detection, scanning-tunneling microscopy (STM-ESR), and miniature microwave scanning probe. Nevertheless all these methods can be employed only to a very limited extent when botanically and biologically-related or relatively thick samples are considered. Thus, for example, the detection by magnetic tips as described in U.S. Pat. No. 6,683,451 suffers from low 3D sensitivity, especially when the samples are thicker than a few microns. Furthermore, this technique requires extreme physical conditions (high vacuum and often low temperatures), and can be employed only after complicated sample preparation. The STM-ESR is a surface (two-dimensional, or 2D) technique capable of handling only solid state samples placed over a conductive surface and also required extreme physical conditions for successful operation. The Hall detection and the miniature microwave scanning probe methods also operates only on the surface, or slightly below it, and have not proven to be useful in micron resolution imaging.
A number of problems in nuclear magnetic resonance imaging have been observed, including the need for strong magnetic fields, requiring expensive superconducting magnets, and the limited spatial resolution of images that are obtained.
There is a need for magnetic resonance imaging systems and methods that provide high resolution 2D and 3D images, especially for thin and thick biologically-related samples, at modest cost, and in short (1-10 min) acquisition times.